Factors causing spatial heterogeneity in soil properties, plant cover, and soil fauna in a non-reclaimed post-mining site

This study explored processes responsible for environmental heterogeneity at a man-made post-mining landscape formed by heaping of homogeneous overburden in longitudinal “waves”. Soil chemistry (pH; conductivity; total C; total, available, and water-soluble P; and available K, Na, and Ca), the distribution of woody and herbaceous plants, litter quantity, and the distribution of soil fauna were studied along a100-m transect. Heterogeneity between waves was very low, and only available P and biomass of herbaceous plants differed among waves. In contrast, all investigated parameters except of total and available P and fauna density differed among position inside waves. Woody plants were more numerous on the wave tops and slopes than on the bottoms. Distribution of shrubs between waves affect between-waves distribution of other parameters, namely soil C_ox, as waves with more shrubs have larger litter input and more soil carbon.     

Chiral imprinted polymers as enantiospecific coatings of stir bar sorptive extraction devices

This paper reports the design of Molecularly Imprinted Polymers (MIP) with affinity towards (S)-citalopram using computational modeling for the selection of functional monomers and monomer:template ratio. Acrylamide was selected as functional monomer and the final complex functional monomer/template resulted in a 3:1 ratio. The polymer was synthesized by radical polymerization initiated by UV onto magnetic stir-bars in order to obtain a stir bar sorptive extraction (SBSE) device capable of selective enantiomeric recognition. After successful template removal, the parameters affecting the SBSE procedure (sample volume, ionic strength, extraction time and pH) were optimized for the effective rebinding of the target analyte. The resultant chirally imprinted polymer based stir-bar was able to selectively extract (S)-citalopram from a racemic mixture in an aqueous media with high specificity (specificity factor 4) between 25 and 500 μgL(-1). The MIP coated stir-bars can have significance for enantiospecific sample pre-concentration and subsequent analysis without the need for any chiral chromatographic separation.     

New reactive polymer for protein immobilisation on sensor surfaces

Immobilisation of biorecognition elements on transducer surfaces is a key step in the development of biosensors. The immobilisation needs to be fast, cheap and most importantly should not affect the biorecognition activity of the immobilised receptor. A novel protocol for the covalent immobilisation of biomolecules containing primary amines using an inexpensive and simple polymer is presented. This tri-dimensional (3D) network leads to a random immobilisation of antibodies on the polymer and ensures the availability of a high percentage of antibody binding sites. The reactivity of the polymer is based on the reaction between primary amines and thioacetal groups included in the polymer network. These functional groups (thioacetal) do not need any further activation in order to react with proteins, making it attractive for sensor fabrication. The novel polymer also contains thiol derivative groups (disulphide groups or thioethers) that promote self-assembling on a metal transducer surface. For demonstration purposes the polymer was immobilised on Au Biacore chips. The resulting polymer layer was characterised using contact angle meter, atomic force microscopy (AFM) and ellipsometry. A general protocol suitable for the immobilisation of bovine serum albumin (BSA), enzymes and antibodies such as polyclonal anti-microcystin-LR antibody and monoclonal anti-prostate specific antigen (anti-PSA) antibody was then optimised. The affinity characteristics of developed immunosensors were investigated in reaction with microcystin-LR, and PSA. The calculated detection limit for analytes depended on the properties of antibodies. The detection limit for microcystin-LR was 10 ngmL(-1) and for PSA 0.01 ngmL(-1). The non-specific binding of analytes to synthesised polymers was very low. The polymer-coated chips were stored for up to 2 months without any noticeable deterioration in their ability to react with proteins. These findings make this new polymer very promising for the development of low-cost, easy to prepare and sensitive biosensors.     

The effect of topsoil removal in restored heathland on soil fauna, topsoil microstructure, and cellulose decomposition: implications for ecosystem restoration

Communities of soil macrofauna, oribatid mites, and nematodes as well as vegetation and soil chemistry were studied on twelve plots representing three replicates of the following treatments: agricultural meadow, heathland, and heathland restored either by partial or complete topsoil removal 15 years earlier. We also studied the effect of soil macrofauna on decomposition and the microstructure of the soil surface layer with litterbags in combination with the analysis of thin soil sections. The communities of soil macrofauna and oribatid mites significantly differed between agricultural meadows and heathlands. The partial and complete topsoil removal plots were more similar to heathlands with respect to macrofauna and to agricultural meadows with respect to oribatid mites. The density and diversity of soil macrofauna was higher in agricultural meadows than in heathlands; in particular, earthworms, litter transformers, root feeders, and microsaprophags were more abundant on meadows. Heathlands, in contrast, contained a much higher diversity of oribatid mites. The community structure of nematodes did not significantly differ among the treatments. Analysis of thin soil sections revealed a thick organic fermentation layer in heathlands, which was absent in agricultural meadows and only weakly developed in the topsoil removal plots. In agricultural meadows, litterbags and thin soil sections indicated that abundant macrofauna, including endogeic earthworms, were very effective in removing the litter from the soil surface and mixing it into the mineral soil. Possible effects of this soil mixing on restoration success are discussed.     

Virtual imprinting as a tool to design efficient MIPs for photosynthesis-inhibiting herbicides

Molecular modelling and computational screening were used to identify functional monomers capable of interacting with several different photosynthesis-inhibiting herbicides. The process involved the design of a virtual library of molecular models of functional monomers containing polymerizable residues and residues able to interact with the template through electrostatic, hydrophobic, Van der Waals forces and dipole-dipole interactions. Each of the entries in the virtual library was probed for its possible interactions with molecular models of the template molecules. It was anticipated that the monomers giving the highest binding score would represent good candidates for the preparation of affinity polymers. Strong interactions were computationally determined between acidic functional monomers like methacrylic acid (MAA) or itaconic acid (IA) with triazines, and between vinylimidazole with bentazone and bromoxynil. Nevertheless, weaker interactions were seen with phenylureas. The corresponding blank polymers were prepared using the selected monomers and tested in the solid phase extraction (SPE) of herbicides from chloroform solutions. A good correlation was found between the binding score of the monomers and the affinities of the corresponding polymers. The use of computationally designed blanks can potentially eliminate the need for molecular imprinting, (adding a template to the monomer mixture to create specific binding sites). Data also showed that some monomers have a natural selectivity for some herbicides, which can be further enhanced by imprinting. Thus, in regard to retention on the blank polymer, we can estimate if the resulting imprinted polymer will be effective or not.     

In silicio search for genes encoding peroxisomal proteins in Saccharomyces cerevisiae

The biogenesis of peroxisomes involves the synthesis of new proteins that after, completion of translation, are targeted to the organelle by virtue of peroxisomal targeting signals (PTS). Two types of PTSs have been well characterized for import of matrix proteins (PTS1 and PTS2). Induction of the genes encoding these matrix proteins takes place in oleate-containing medium and is mediated via an oleate response element (ORE) present in the region preceding these genes. The authors have searched the yeast genome for OREs preceding open reading frames (ORFs), and for ORFs that contain either a PTS1 or PTS2. Of the ORFs containing an ORE, as well as either a PTS1 or a PTS2, many were known to encode bona fide peroxisomal matrix proteins. In addition, candidate genes were identified as encoding putative new peroxisomal proteins. For one case, subcellular location studies validated the in silicio prediction. This gene encodes a new peroxisomal thioesterase.     

Nonaprenyl-4-hydroxybenzoate transferase, an enzyme involved in ubiquinone biosynthesis, in the endoplasmic reticulum-Golgi system of rat liver

The properties and distribution of nonaprenyl-4-hydroxybenzoate transferase in rat liver were investigated with subcellular fractions, liver perfusion, and in vivo labeling with [3H]solanesyl-PP. In addition to some ubiquinone-9, only one labeled intermediate, i.e. nonaprenyl-4-hydroxybenzoate, was obtained. In the total microsomal fraction, the enzyme had a pH optimum of 7.5 and was completely inhibited by Triton X-100 and deoxycholate, but not by taurodeoxycholate and beta-octyl glucoside. Liver, kidney, and spleen demonstrated the highest activities of nonaprenyl-4-hydroxybenzoate transferase. Upon subcellular fractionation, high specific activities were found in smooth II microsomes and Golgi III vesicles. The enzyme was also found in lysosomes and plasma membranes, but only at low levels in rough and smooth I microsomes and mitochondria and not at all in peroxisomes and cytosol. When the product of the transferase reaction was used as a substrate in vitro and in a perfusion system, the only product obtained was end product ubiquinone-9. Although the transferase reaction was associated with the inner, luminal surface of microsomal vesicles, the terminal reaction(s) for ubiquinone-9 synthesis are found at the outer cytoplasmic surface. The results suggest that the major site for ubiquinone synthesis is the endoplasmic reticulum-Golgi system, which also participates in the distribution of ubiquinone-9 to other cellular membranes.     

Suppression of NB-LRR genes by miRNAs promotes nitrogen-fixing nodule development in Medicago truncatula

Plant genomes contain two major classes of innate immune receptors to recognize different pathogens. The pattern recognition receptors perceive conserved pathogen-associated molecular patterns and the resistance genes with nucleotide-binding (NB) and leucine-rich repeat (LRR) domains recognize specific pathogen effectors. The precise regulation of resistance genes is important since the unregulated expression of NB-LRR genes can inhibit growth and may result in autoimmunity in the absence of pathogen infection. It was shown that a subset of miRNAs could target NB-LRR genes and act as an important regulator of plant immunity in the absence of pathogens. Plants not only interact with pathogens, but they can also establish symbiotic interactions with microbes. Nitrogen-fixing symbiotic interaction and nodule formation of legumes may also require the suppression of host defence to prevent immune responses. We found that upon symbiotic interactions, miRNAs repressing NB-LRR expression are upregulated in the developing nodules of Medicago truncatula. Furthermore, we show that the suppression of the activity of the NB-LRR genes targeted by these miRNAs is important during nodule development. Our results suggest that the downregulation of NB-LRR resistance genes in the developing nodule produces a suitable niche that facilitates bacterial colonization and the development of an N-fixing nodule.